Distribution of methotrexate (MTX), citrovorum factor (CF), and 5-methyltetrahydrofolate (5-MTHFA) will be followed in serum and cerebrospinal fluid of patients with osteogenic sarcoma and meningeal carcinomatosis, using microbiological as well as other assay methods, viz. high pressure liquid chromatography, radioimmunoassay, "Emit-and" enzyme immunoassay, and enzymic assay methods, developed in our laboratory. We propose to establish guidelines for toxicity in relation to serum concentrations of MTX, CF and 5-MTHFA, as well as to determine the effectiveness of the therapy protocol. Distribution kinetics of MTX, CF and 5-MTHFA will also be followed in brain tumor models in rats and monkeys established by Dr. Shapiro and associates. We also propose to isolate and characterize antibiotic resistant strains of organisms used for the assay of MTX, CF, 5-MTHFA and examine these strains for the physiological alterations associated with the newly acquired resistance.